Synchronised monitoring of plant and insect diversity: a case study using automated Malaise traps and DNA-based methods.

The occurrence and distribution of insects and their possible associations with plant species are largely unknown in Germany and baseline data to monitor future trends are urgently needed. Using newly-designed automated Malaise trap multi-samplers, the occurrence of insect species and their potential associations with plants was monitored synchronously at two contrasting field sites in Germany: an urban botanical garden and a forest research station. Taxa were identified by metabarcoding of the insects and the plant traces present in the preservative ethanol of the Malaise trap samples. For comparison, a botanical survey was conducted in the vicinity of the traps. Across both sites, we identified a total of 1290 exact sequence variants (ESVs) assigned to Insecta, of which 205 are known to be pollinators. In the botanical garden, we detected the occurrence of 128 plant taxa, of which 41 also had one of their known insect pollinator species detected. Insect species richness was highest in May, mainly attributed to an increase in Diptera. These results present a case study of the applicability of automated sampling and DNA-based methods to monitor the timings of flowering and corresponding activity of plant-visiting insects.

Biodiversity and Spatiotemporal Variations of Mecoptera in Thailand: Influences of Elevation and Climatic Factors.

Ecological analyses of the small and lesser-known insect order Mecoptera in Thailand are presented. Specimens were collected monthly over a period of 12 consecutive months, using both Malaise and pan traps, from 29 sampling sites located in 18 national parks throughout Thailand. A total of 21 species in four genera were identified from 797 specimens, including Panorpa (1 species), Neopanorpa (18 species), Bittacus (1 species), and Terrobittacus (1 species), with the latter genus representing a new genus record to Thailand. Neopanorpa harmandi, N. siamensis, N. byersi, and N. malaisei were the most abundant species, representing 27.4%, 11.3%, 10.3% and 8.8% of the total specimens, respectively. The species with the highest frequency, as indicated by the high percentage of species occurrence (%SO), was N. siamensis (51%), followed by N. byersi (34%), N. harmandi (34%), N. spatulata (27%), and N. inchoata (27%). Eleven species (52%) exhibited specific regional occurrences. N. tuberosa and N. siamensis had the widest distribution, being found in almost all regions except for western and southern regions for the first and second species, respectively. The seasonal species richness of Mecoptera was high during the rainy season in the northern, northeastern, central, eastern, and western regions, with the highest richness observed in July (15 species), followed by the hot (10 species) and cold seasons (7 species), while there was no significant difference in species richness between seasons in the southern region. Multiple regression models revealed a negative association between species richness and abundance of Mecoptera with both elevation and temperature, and a positive association between rainfall and species evenness. It is predicted that climatic changes will have a detrimental effect on the mecopteran community. The results of this study enhance the understanding of the ecological aspects of Mecoptera, offering crucial insights into its biodiversity and distribution, which are vital for conservation and forest management.

Four new species of Sphaeroderma Stephens (Coleoptera, Chrysomelidae, Galerucinae, Alticini) from Taiwan, with discussion on genus boundaries based on S.flavonotatum Chûjô and S.jungchani sp. nov.

Three new species of Sphaeroderma Stephens, 1831, S.hsuisp. nov., S.changisp. nov., and S.sheipaensissp. nov. are described based on specimens from Shei-Pa National Park, Taiwan. A fourth new species, S.jungchanisp. nov., is described based on specimens from southern Taiwan. Delimitation of the genus is discussed based on S.flavonotatum Chûjô, 1937, which is redescribed, and the new species, S.jungchanisp. nov.

Optimizing insect metabarcoding using replicated mock communities.

1: Metabarcoding (high-throughput sequencing of marker gene amplicons) has emerged as a promising and cost-effective method for characterizing insect community samples. Yet, the methodology varies greatly among studies and its performance has not been systematically evaluated to date. In particular, it is unclear how accurately metabarcoding can resolve species communities in terms of presence-absence, abundances, and biomass. 2: Here we use mock community experiments and a simple probabilistic model to evaluate the effect of different DNA extraction protocols on metabarcoding performance. Specifically, we ask four questions: (Q1) How consistent are the recovered community profiles across replicate mock communities?; (Q2) How does the choice of lysis buffer affect the recovery of the original community?; (Q3) How are community estimates affected by differing lysis times and homogenization?; and (Q4) Is it possible to obtain adequate species abundance estimates through the use of biological spike-ins? 3: We show that estimates are quite variable across community replicates. In general, a mild lysis protocol is better at reconstructing species lists and approximate counts, while homogenization is better at retrieving biomass composition. Small insects are more likely to be detected in lysates, while some tough species require homogenization to be detected. Results are less consistent across biological replicates for lysates than for homogenates. Some species are associated with strong PCR amplification bias, which complicates the reconstruction of species counts. Yet, with adequate spike-in data, species abundance can be determined with roughly 40% standard error for homogenates, and with roughly 50% standard error for lysates, under ideal conditions. In the latter case, however, this often requires species-specific reference data, while spike-in data generalizes better across species for homogenates. 4: We conclude that a non-destructive, mild lysis approach shows the highest promise for presence/absence description of the community, while also allowing future morphological or molecular work on the material. However, homogenization protocols perform better for characterizing community composition, in particular in terms of biomass.

DNA barcoding data release for the Phoridae (Insecta, Diptera) of the Halimun-Salak National Park (Java, Indonesia).

Launched in 2015, the large-scale initiative Indonesian Biodiversity Discovery and Information System (IndoBioSys) is a multidisciplinary German-Indonesian collaboration with the main goal of establishing a standardised framework for species discovery and all associated steps. One aspect of the project includes the application of DNA barcoding for species identification and biodiversity assessments. In this framework, we conducted a large-scale assessment of the insect fauna of the Mount Halimun-Salak National Park which is one of the largest tropical rain-forest ecosystems left in West Java. In this study, we present the results of processing 5,034 specimens of Phoridae (scuttle flies) via DNA barcoding. Despite limited sequencing success, we obtained more than 500 clusters using different algorithms (RESL, ASAP, SpeciesIdentifier). Moreover, Chao statistics indicated that we drastically undersampled all trap sites, implying that the true diversity of Phoridae is, in fact, much higher. With this data release, we hope to shed some light on the hidden diversity of this megadiverse group of flies.

Robber flies and hover flies (Insecta, Diptera, Asilidae and Syrphidae) in beech forests of the central Apennines: a contribution to the inventory of insect biodiversity in Italian State Nature Reserves.

Background: The present paper describes a sampling-event dataset on species belonging to two families of Diptera (Syrphidae and Asilidae) collected between 2012 and 2019 in two Italian beech forests located in the central Apennines. The reference dataset consists of an annotated checklist and has been published on Zenodo. Syrphidae and Asilidae are two widespread and key ecological groups, including predator, pollinator and saproxylic species. Despite their pivotal role in both natural and man-made ecosystems, these families are still poorly known in terms of local distribution and open-access sampling-event data are rare in Italy. New information: This open-access dataset includes 2,295 specimens for a total of 21 Asilidae and 65 Syrphidae species. Information about the collection (e.g. place, date, methods applied, collector) and the identification (e.g. species name, author, taxon ID) of the species is provided. Given the current biodiversity crisis, the publication of checklists, sampling-event data and datasets on insect communities in open-access repositories is highly recommended, as it represents the opportunity to share biodiversity information amongst different stakeholders. Moreover, such data are also a valuable source of information for nature reserve managers responsible for monitoring the conservation status of protected and endangered species and habitats and for evaluating the effects of conservation actions over time.

Extensive regional variation in the phenology of insects and their response to temperature across North America.

Climate change models often assume similar responses to temperatures across the range of a species, but local adaptation or phenotypic plasticity can lead plants and animals to respond differently to temperature in different parts of their range. To date, there have been few tests of this assumption at the scale of continents, so it is unclear if this is a large-scale problem. Here, we examined the assumption that insect taxa show similar responses to temperature at 96 sites in grassy habitats across North America. We sampled insects with Malaise traps during 2019-2021 (N = 1041 samples) and examined the biomass of insects in relation to temperature and time of season. Our samples mostly contained Diptera (33%), Lepidoptera (19%), Hymenoptera (18%), and Coleoptera (10%). We found strong regional differences in the phenology of insects and their response to temperature, even within the same taxonomic group, habitat type, and time of season. For example, the biomass of nematoceran flies increased across the season in the central part of the continent, but it only showed a small increase in the Northeast and a seasonal decline in the Southeast and West. At a smaller scale, insect biomass at different traps operating on the same days was correlated up to ~75 km apart. Large-scale geographic and phenological variation in insect biomass and abundance has not been studied well, and it is a major source of controversy in previous analyses of insect declines that have aggregated studies from different locations and time periods. Our study illustrates that large-scale predictions about changes in insect populations, and their causes, will need to incorporate regional and taxonomic differences in the response to temperature.

Contributions of CO2-Baited Malaise-Type Traps to the Knowledge of Hematophagous and Oestrid Flies (Diptera: Oestridae) Parasitizing Cervid Hosts.

This review describes an innovative and efficient modification of a type of Malaise trap meme overlooked in a recent review of such insect traps. It further identifies the large variety of dipteran parasites of vertebrates caught in the traps when CO2 was added as an attractant that otherwise rarely were caught in unbaited traps. Baited trap catches of parasitic flies mimicked those caught attacking cervid hosts. This review particularly focuses on comprehensive studies of: 1) several hematophagous species of largely unknown snipe flies (Diptera: Rhagionidae: Symphoromyia [Artiodactyla: Cervidae]) found host specific for Columbian black-tailed deer (Odocoileus hemionus columbianus Richardson), and 2) the biology of oestrid fly (Diptera: Oestridae) parasites of black-tailed deer and caribou/reindeer [Rangifer tarandus (L.) (Artiodactyla: Cervidae)], after it was discovered that the non-hematophagous females are guided to their vertebrate hosts by tracking a source of CO2, as done by many hematophagous flies.

Using Malaise Traps and Metabarcoding for Biodiversity Assessment in Vineyards: Effects of Weather and Trapping Effort.

Metabarcoding is a powerful tool for ecological studies and monitoring that might provide a solution to the time-consuming taxonomic identification of the vast diversity of insects. Here, we assess how ambient weather conditions during Malaise trap exposure and the effort of trapping affect biomass and taxa richness in vineyards. Biomass varied by more than twofold with weather conditions. It increased with warmer and drier weather but was not significantly related with wind or precipitation. Taxa richness showed a saturating relationship with increasing trapping duration and was influenced by environmental and seasonal effects. Taxa accumulation was high, increasing fourfold from three days of monthly trap exposure compared to continuous trapping and nearly sixfold from sampling at a single site compared to 32 sites. The limited saturation was mainly due to a large number of singletons, such as rare species, in the metabarcoding dataset. Metabarcoding can be key for long-term insect monitoring. We conclude that single traps operated for up to ten days per month are suitable to monitor the presence of common species. However, more intensive trapping is necessary for a good representation of rare species in biodiversity monitoring. The data collected here can potentially guide the design of monitoring studies.

Metabarcoding reveals massive species diversity of Diptera in a subtropical ecosystem.

Diptera is often considered to be the richest insect group due to its great species diversity and broad ecological versatility. However, data on dipteran diversity from subtropical ecosystems have hitherto been scarce, due to the lack of studies conducted at an appropriate large scale. We investigated the diversity and composition of Diptera communities on Tianmu Mountain, Zhejiang, China, using DNA metabarcoding technology, and evaluated their dynamic responses to the effects of slope aspect, season, and altitudinal zone. A total of 5,092 operational taxonomic units (OTUs) were discovered and tentatively assigned to 72 dipteran families, including 2 family records new for China and 30 family records new for the locality. Cecidomyiidae, Sciaridae, and Phoridae were the predominant families, representing 53.6% of total OTUs, while 52 families include >95% unidentified and presumed undescribed species. We found that the community structure of Diptera was significantly affected by aspect, seasonality (month) and elevation, with richer diversity harbored in north-facing than south-facing slopes, and seasonality a more profound driver of community structure and diversity than elevation. Overall, massive species diversity of Diptera communities was discovered in this subtropical ecosystem of east China. The huge diversity of potentially undescribed species only revealed by metabarcoding now requires more detailed taxonomic study, as a step toward an evolutionary integration that accumulates information on species' geographic ranges, ecological traits, functional roles, and species interactions, and thus places the local communities in the context of the growing knowledge base of global biodiversity and its response to environmental change.

Community Structure, Biodiversity and Spatiotemporal Distribution of the Black Flies (Diptera: Simuliidae) Using Malaise Traps on the Highest Mountain in Thailand.

Black flies form a group of small blood-sucking insects of medical and veterinary importance. This study aimed to investigate the community structure, biodiversity and spatial and temporal distribution of adult black flies in tropical rain forests, by using malaise traps in Doi Inthanon National Park, northern Thailand. Malaise traps were placed along six elevational gradients (400 m to 2500 m, above sea level) at Doi Inthanon National Park, Chiang Mai province, from December 2013 to November 2014. A total of 9406 adult female black flies belonging to five subgenera-Daviesellum (2%), Gomphostilbia (23%), Montisimulium (11%), Nevermannia (16%) and Simulium (48%)-were collected. Among 44 taxa found, S. tenebrosum complex had the highest relative abundance (11.1%), followed by the S. asakoae species-group (9.6%), the S. striatum species-group (7.7%), S. inthanonense (6.6%), S. doipuiense complex (6.4%), S. chomthongense complex (5.3%), S. chumpornense (5.1%) and S. nigrogilvum (4.1%). Two human-biting species-S. nigrogilvum and species in the S. asakoae species-group-were found in all of the collection sites with 100% species occurrence. Species richness was highest at mid elevation (1400 m), which is represented by 19 black fly species. The peak and lowest seasonal abundance was observed in the rainy and hot season, respectively. Seasonal species richness was highest in the cold season, except for that from elevation sites at 700 m, 1700 m and 2500 m. This study revealed that the malaise trap is effective in providing important data for further monitoring of the effects of environmental changes and conservation planning on the biodiversity of black flies in Doi Inthanon National Park.

The data of the Swedish Malaise Trap Project, a countrywide inventory of Sweden's insect fauna.

BACKGROUND: Despite Sweden's strong entomological tradition, large portions of its insect fauna remain poorly known. As part of the Swedish Taxonomy Initiative, launched in 2002 to document all multi-cellular species occurring in the country, the first taxonomically-broad inventory of the country's insect fauna was initiated, the Swedish Malaise Trap Project (SMTP). In total, 73 Malaise traps were deployed at 55 localities representing a wide range of habitats across the country. Most traps were run continuously from 2003 to 2006 or for a substantial part of that time period. The total catch is estimated to contain 20 million insects, distributed over 1,919 samples (Karlsson et al. 2020). The samples have been sorted into more than 300 taxonomic units, which are made available for expert identification. Thus far, more than 100 taxonomists have been involved in identifying the sorted material, recording the presence of 4,000 species. One third of these had not been recorded from Sweden before and 700 have tentatively been identified as new to science. NEW INFORMATION: Here, we describe the SMTP dataset, published through the Global Biodiversity Information Facility (GBIF). Data on the sorted material are available in the "SMTP Collection Inventory" dataset. It currently includes more than 130,000 records of taxonomically-sorted samples. Data on the identified material are published using the Darwin Core standard for sample-based data. That information is divided up into group-specific datasets, as the sample set processed for each group is different and in most cases non-overlapping. The current data are divided into 79 taxonomic datasets, largely corresponding to taxonomic sorting fractions. The orders Diptera and Hymenoptera together comprise about 90% of the specimens in the material and these orders are mainly sorted to family or subfamily. The remaining insect taxa are mostly sorted to the order level. In total, the 79 datasets currently available comprise around 165,000 specimens, that is, about 1% of the total catch. However, the data are now accumulating rapidly and will be published continuously. The SMTP dataset is unique in that it contains a large proportion of data on previously poorly-known taxa in the Diptera and Hymenoptera.

Taiwanoshaira Lee & Beenen, a new genus and first record of moss-inhabiting Galerucinaesensu stricto (Coleoptera, Chrysomelidae) from Taiwan.

Taiwanoshaira Lee & Beenen gen. nov. is described. It represents the first genus of Galerucinae (s. str.) in Taiwan documented to inhabit moss cushions. Shaira chujoi Kimoto, 1982 is transferred to Taiwanoshaira, as follows: T. chujoi (Kimoto), comb. nov. Two new species, T. taipingshanensis sp. nov. and T. tsoui sp. nov., are described. Adults of T. taipingshanensis sp. nov. were observed feeding on the moss species Plagiomnium vesicatum (Besch.) T.J. Kop. (Mniaceae). Microhabitats and distribution of Taiwanoshaira species are discussed.

Toward a standardized quantitative and qualitative insect monitoring scheme.

The number of insect species and insect abundances decreased severely during the past decades over major parts of Central Europe. Previous studies documented declines of species richness, abundances, shifts in species composition, and decreasing biomass of flying insects. In this study, we present a standardized approach to quantitatively and qualitatively assess insect diversity, biomass, and the abundance of taxa, in parallel. We applied two methods: Malaise traps, and automated and active light trapping. Sampling was conducted from April to October 2018 in southern Germany, at four sites representing conventional and organic farming. Bulk samples obtained from Malaise traps were further analyzed using DNA metabarcoding. Larger moths (Macroheterocera) collected with light trapping were further classified according to their degree of endangerment. Our methods provide valuable quantitative and qualitative data. Our results indicate more biomass and higher species richness, as well as twice the number of Red List lepidopterans in organic farmland than in conventional farmland. This combination of sampling methods with subsequent DNA metabarcoding and assignments of individuals according depending on ecological characteristics and the degree of endangerment allows to evaluate the status of landscapes and represents a suitable setup for large-scale long-term insect monitoring across Central Europe, and elsewhere.

The Swedish Malaise Trap Project: A 15 Year Retrospective on a Countrywide Insect Inventory.

The Swedish Malaise Trap Project (SMTP) is one of the most ambitious insect inventories ever attempted. The project was designed to target poorly known insect groups across a diverse range of habitats in Sweden. The field campaign involved the deployment of 73 Malaise traps at 55 localities across the country for three years (2003-2006). Over the past 15 years, the collected material has been hand sorted by trained technicians into over 300 taxonomic fractions suitable for expert attention. The resulting collection is a tremendous asset for entomologists around the world, especially as we now face a desperate need for baseline data to evaluate phenomena like insect decline and climate change. Here, we describe the history, organisation, methodology and logistics of the SMTP, focusing on the rationale for the decisions taken and the lessons learned along the way. The SMTP represents one of the early instances of community science applied to large-scale inventory work, with a heavy reliance on volunteers in both the field and the laboratory. We give estimates of both staff effort and volunteer effort involved. The project has been funded by the Swedish Taxonomy Initiative; in total, the inventory has cost less than 30 million SEK (approximately 3.1 million USD). Based on a subset of the samples, we characterise the size and taxonomic composition of the SMTP material. Several different extrapolation methods suggest that the material comprises around 20 million specimens in total. The material is dominated by Diptera (75% of the specimens) and Hymenoptera (15% of specimens). Amongst the Diptera, the dominant groups are Chironomidae (37% of specimens), Sciaridae (15%), Phoridae (13%), Cecidomyiidae (9.5%) and Mycetophilidae (9.4%). Within Hymenoptera, the major groups are Ichneumonidae (44% of specimens), Diaprioidea (19%), Braconidae (9.6%), Platygastroidea (8.5%) and Chalcidoidea (7.9%). The taxonomic composition varies with latitude and season. Several Diptera and Hymenoptera groups are more common in non-summer samples (collected from September to April) and in the North, while others show the opposite pattern. About 1% of the total material has been processed and identified by experts so far. This material represents over 4,000 species. One third of these had not been recorded from Sweden before and almost 700 of them are new to science. These results reveal the large amounts of taxonomic work still needed on Palaearctic insect faunas. Based on the SMTP experiences, we discuss aspects of planning and conducting future large-scale insect inventory projects using mainly traditional approaches in relation to more recent approaches that rely on molecular techniques.

Using Malaise traps for collecting Lepidoptera (Insecta), with notes on the preparation of Macrolepidoptera from ethanol.

The present paper deals with the potential of employing Malaise traps for collecting butterflies and moths for morphological analysis and presents a protocol for preparing Macrolepidoptera from Malaise trap samples that were preserved in ethanol. About 80 specimens of Lepidoptera, including Nymphalidae, Geometridae, Hesperiidae, Erebidae, Noctuidae, Pyralidae and Tortricidae, were mounted, following the protocol. All specimens with robust wings and contrasting wing patterns were well suited for the study of external morphology, regardless of the family. The specimens used in this study were collected in highland forest areas of central Vietnam with a little known entomofauna, as part of the German-Vietnamese biodiversity project 'VIETBIO'. The study offers new methodological approaches in an attempt to make the most of the material that was obtained using Malaise traps.

DNA barcodes reveal deeply neglected diversity and numerous invasions of micromoths in Madagascar 1.

Madagascar is a prime evolutionary hotspot globally, but its unique biodiversity is under threat, essentially from anthropogenic disturbance. There is a race against time to describe and protect the Madagascan endangered biota. Here we present a first molecular characterization of the micromoth fauna of Madagascar. We collected 1572 micromoths mainly using light traps in both natural and anthropogenically disturbed habitats in 24 localities across eastern and northwest Madagascar. We also collected 1384 specimens using a Malaise trap in a primary rain forest at Andasibe, eastern Madagascar. In total, we DNA barcoded 2956 specimens belonging to 1537 Barcode Index Numbers (BINs), 88.4% of which are new to BOLD. Only 1.7% of new BINs were assigned to species. Of 47 different families found, Dryadaulidae, Bucculatricidae, Bedelliidae, Batrachedridae, and Blastobasidae are newly reported for Madagascar and the recently recognized Tonzidae is confirmed. For test faunas of Canada and Australia, 98.9%-99.4% of Macroheterocera BINs exhibited the molecular synapomorphy of a phenylalanine in the 177th complete DNA barcode codon. Non-macroheteroceran BINs could thus be sifted out efficiently in the Malaise sample. The Madagascar micromoth fauna shows highest affinity with the Afrotropics (146 BINs also occur in the African continent). We found 22 recognised pests or invasive species, mostly occurring in disturbed habitats. Malaise trap samples show high temporal turnover and alpha diversity with as many as 507 BINs collected; of these, astonishingly, 499 (98.4%) were novel to BOLD and 292 (57.6%) were singletons. Our results provide a baseline for future surveys across the island.

Expedited assessment of terrestrial arthropod diversity by coupling Malaise traps with DNA barcoding 1.

Monitoring changes in terrestrial arthropod communities over space and time requires a dramatic increase in the speed and accuracy of processing samples that cannot be achieved with morphological approaches. The combination of DNA barcoding and Malaise traps allows expedited, comprehensive inventories of species abundance whose cost will rapidly decline as high-throughput sequencing technologies advance. Aside from detailing protocols from specimen sorting to data release, this paper describes their use in a survey of arthropod diversity in a national park that examined 21 194 specimens representing 2255 species. These protocols can support arthropod monitoring programs at regional, national, and continental scales.

Comparisons of Citizen Science Data-Gathering Approaches to Evaluate Urban Butterfly Diversity.

By 2030, ten percent of earth's landmass will be occupied by cities. Urban environments can be home to many plants and animals, but surveying and estimating biodiversity in these spaces is complicated by a heterogeneous built environment where access and landscaping are highly variable due to human activity. Citizen science approaches may be the best way to assess urban biodiversity, but little is known about their relative effectiveness and efficiency. Here, we compare three techniques for acquiring data on butterfly (Lepidoptera: Rhopalocera) species richness: trained volunteer Pollard walks, Malaise trapping with expert identification, and crowd-sourced iNaturalist observations. A total of 30 butterfly species were observed; 27 (90%) were recorded by Pollard walk observers, 18 (60%) were found in Malaise traps, and 22 (73%) were reported by iNaturalist observers. Pollard walks reported the highest butterfly species richness, followed by iNaturalist and then Malaise traps during the four-month time period. Pollard walks also had significantly higher species diversity than Malaise traps.

New record of the genus Zombrus Marshall, 1897 (Hymenoptera: Braconidae: Doryctinae), with description of three new species from Vietnam.

The genus Zombrus Marshall, 1897, is newly recorded for Vietnam with four species. Three new species (Z. bicoloricorpus Long, sp. n., Z. flavicorpus Long van Achterberg, sp. n., and Z. tricoloricorpus Long, sp. n.) are described and illustrated, and one species (Z. pallidiventris Cameron, 1910) is reported for the first time from Vietnam and outside Indonesia. The key to Oriental and Palaearctic species of Zombrus and a checklist with distributions of the nine species from Oriental and Palaearctic regions are provided.